Characterization Of T2 To T4 Transgenic Lines Derived From Double Resistance Gene For Rice Blast Via Multigene Assembly And Transformation Vector System

Rice(Oryza sativa)is the main food for more than 50% of the world’s population.Rice blast,caused by the filamentous ascomycete fungus Magnaporthe oryzae,is one of the most destructive rice diseases in the world,which seriously affects rice production.Research and practice at home and abroad have proven that the development and use of disease resistance genes(R)is the most economical,environmentally friendly and effective measure to control rice blast.However,due to the genetic complexity and mutability of rice blast fungus,the resistance of most resistant varieties to rice blast will be weakened or even lost after 3 to 5 years of cultivation.Therefore,aggregation of disease resistance genes is considered to be one of the most reliable methods to obtain durable resistance to rice blast.The aggregation of disease resistance genes can select multiple disease resistance genes at the same time and aggregate them into the same variety,thereby improving resistance and broadening the resistance spectrum to obtain durable resistance.In the previous research in this laboratory,through the genetic transformation of double gene has obtained the Pik/Pi1(43),the Pik/Pikh(43),the Pik/Pi36(43),Pi7/Pi1(43),Pi7/Pikh(43),Pi7/Pi36(43)combination of the six transgenic T1 sgeneration seedlings and their seeds.The purpose of this study is to screen out resistant plants that do not contain selection markers and contain complete double genes;compare the resistance between different combinations and study the effects of different combinations on rice resistance.This study carried out the following work,the results are as follows:1.Phenotype identification of anti-susceptibility separationAmong the 838 T2-T4 strains with 6 double-gene combinations,37 strains with a single gene segregation ratio were identified,that is,the resistance ratio was 3:1,and a total of 45 fully resistant strains were identified.Among them,Pik/Pi1(43)has 6 strains whose resistance to susceptibility ratio is in line with the single gene segregation ratio,6 strains are fully resistant strains;Pik/Pikh(43)has 6 strains and 12 strains respectively Lines;Pik/Pi36(43)has 3 lines and 9 lines respectively;Pi7/Pi1(43)has 10 lines and 6 lines respectively;Pi7/Pikh(43)has 3 lines respectively And 11 lines;Pi7/Pi36(43)has 9 lines and1 line respectively.2 Molecular identification of selection markers and double genes341 T2 generation test individuals,314 T3 generation test individuals,and 185 T4 generation test individuals of 6 dual gene combinations were selected for specific molecular identification of disease resistance genes.The results showed that the combination of Pik/Pi1(43)obtained 30 resistant plants with no selectable markers and double genes;Pik/Pikh(43)obtained 41 plants,Pi7/Pi1(43)obtained 34 plants,Pi7/Pikh(43)Obtained 32 plants,but Pik/Pi36(43)and Pi7/Pi36(43)have not been screened for dual gene plants without selection markers.